Nongenomic action and mechanism of 17β-estradiol in cytosolic calcium concentration in delayed implantation mouse endometrial stromal cells

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Author:
WANG Qiang(Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China)
YUE Li-Min(Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China)
ZHANG Jin-Hu(Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China)
TIAN Ji-Mei(Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China)
HE Ya-Ping(Department of Physiology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China)
Journal Title:
ACTA PHYSIOLOGICA SINICA
Issue:
Volume 60, Issue 02, 2008
DOI:
Key Word:
estrogen; phospholipase C; calcium; stromal cells

Abstract: To investigate the existence of nongenomic action of 17β-estradiol (E2) in the delayed implantation mouse endometrialstromal ceils, the changes in intraceilular calcium concentration ([Ca2+]I) and the upstream of calcium signal in vitro were detected. Theexperiment was composed of two parts. Firstly, the change in [Ca2+]I in endometrial stromal cells induced by E2 under differentconditions was detected. The mice were divided into 6 groups as follows: 0.1% dimethylsulfoxide (DMSO) control group, 1×10-8 mol/Lbovine serum albumin (BSA) control group, 1×10-8 mol/L E2 group, 1×10-8 mol/L E2 conjugated with BSA (E2-BSA) group, 1×10-8 mol/LE2 + calcium-free medium group, 1×10-8 mol/L E2 + 5μg/mL tamoxifen group, with 4 mice in each group. The endometrial tissue wasobtained from delayed implantation mice at pregnant day 7, and digested by incubation of tissue minces in Hank's balanced salts(HBSS, pH 7.2), which contained glucose (1 g/L), and eollagenase I (0.125%), for 1 h at 37 ℃. The stromal cells were preloaded with2.5 mmol/L Fluo-3/AM, a fluorescent probe of calcium, for 30 rain. A confocal laser scanning microscope, which fixed the wave lengthof excitation and emission at 488 nm and 526 nm, respectively, was used to detect the change in [Ca2+]I Secondly, the mechanism of E2effects in endometrial stromal cells was investigated. Immunofluorescent method was used to detect the change in phosphorylation ofphospholipase C (PLC) before and after the stromal cells were treated with E2 for 5 rain, 15 rain, and 30 rain. Seven delayedimplantation mice were used. The results were as follows. [Ca2+]I increased immediately and reached the maximum at 15 rain after the stromal cells were treated with 1×10-8 mol/L E2 and returned to the normal level at 30 min. In E2-BSA group and E2 + calcium-free medium group the same results were obtained as that in E2 group, but there was no increase of [Ca2+]I in DMSO and BSA groups. Tamoxifen, a traditional antagonist of estrogen receptor, did not inhibit the increase in [Ca2+]I induced by E2. Immunofluorescent results showed that the change in phosphorylated-PLC level had the same trend as [Ca2+]I after the cells were treated with E2. Compared with that in the control group, the immunofluorescent intensity increased at the beginning and achieved the maximum at 15 min (P<0.001), then declined to the normal level at 30 min. These results suggest that the rapid response of [Ca2+]I induced by E2 in the endometrial stromal cells in delayed implantation mice is possibly carded out through a nongenomic pathway, and the transmembrane signal transduction is related to the phosphorylation of PLC in this process.

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